Respiratory syncytial virus (RSV) is a major etiologic agent of lower respiratory tract infection in children and the elderly. In the U.S., RSV can be detected in 43% of pediatric patients hospitalized with bronchiolitis and 25% with pneumonia. Neonates have well-defined differentially regulated features in adaptive immune responses, and they depend largely on maternal antibodies and innate immune responses for protection against pathogens. Data suggests that neonatal innate responses are immature and exhibit a Th2 biased cytokine response, impaired production of interferon alpha (IFNa), and decreased expression of Toll like receptors (TLRs). Studies suggest that cytokine responses may correlate with disease severity. Furthermore, it has been suggested that cytokine responses, specifically typ I IFNs, to hepatitis C virus may activate expression of cellular microRNAs which exhibit anti-viral activities. The long term goal of this project is to study human immune responses against RSV. The objective of this application is to examine cytokine responses to RSV infection ex vivo. The central hypothesis is that cells of the human immune system respond to RSV through specific cytokine responses, which may determine viral clearance, disease severity, and outcome of infection. To test the central hypothesis, two specific aims will be pursued: 1. Examine differences in DC responses to RSV infection in peripheral blood (PB) vs. cord blood (CB). DCs will be directly isolated from PB and CB using magnetic sorting, will be co-cultured with RSV, and transcription of cytokines will be examined by real-time RT PCR and microarray analysis. Global differences in the transcriptional profiles of PB and CBDCs will be compared using Ingenuity Pathway Analysis software. Differences in the transforming growth factor beta family and related genes will be specifically examined. 2. MicroRNA responses to RSV infection will be studied. Peripheral blood mononuclear cells from adults will be isolated, cocultured with RSV and will be used for microRNA array analysis. Results will be confirmed by real time PCR and mechanisms of activation, with specific interest in cytokine induction of microRNAs will be studied. Public Health Relevance: Respiratory syncytial virus is a significant human pathogen that causes severe bronchiolitis and pneumonia in infants and the elderly. In this project, immune responses to the virus will be examined. Understanding how the immune system responds differently to virus in infants vs. adults may help us understand why infants experience more severe disease and inform efforts to design new vaccines.)